The goal of this project is to address how the parasexual cycle of the fungus Candida albicans influences cell fitness and pathogenesis. This work is part of a long-range goal to understand the role of parasexuality in C. albicans colonization and disease in the mammalian host. C. albicans is the primary cause of oropharyngeal candidiasis (OPC), a condition that continues to afflict HIV-infected patients even in the era of antiretroviral therapy. C. albicans is also a prevalent cause of life-threatening systemic infections, particularly in nosocomial and immunocompromised populations. A defining feature of C. albicans biology is its ability to colonize and infect many sites throughout the human body. This remarkable ability is dependent on genetic and epigenetic mechanisms that act to promote phenotypic plasticity. Strains undergo switching between alternative cell states, and these transitions are essential to C. albicans's lifestyle both as a commensal and as an opportunistic pathogen. Morphologic and phenotypic changes influence virulence, tissue specificity, interactions with immune cells, and entry into the parasexual cycle. It is therefore essential to understand the mechanisms promoting phenotypic variation, and the consequences of such variation for infection of the host. A major goal of this proposal is to define the abilit of the parasexual cycle to generate novel C. albicans strains with the potential to promote pathogenesis. The parasexual cycle involves phenotypic switching to a mating-competent state, cell fusion to generate tetraploid cells, and concerted chromosome loss to form recombinant diploid or aneuploid strains. Exciting preliminary experiments establish that parasexuality can generate strains with increased virulence as well as increased resistance to environmental stress. A systematic analysis of parasex progeny will now be performed to identify isolates with stress-resistant phenotypes including resistance to antifungal drugs, as well as isolates that exhibit increased virulence. Genotyping of these isolates will determine the chromosomal changes that underlie these adaptive events. In particular, it is expected that the presence of specific aneuploid chromosomes will be associated with particular C. albicans phenotypes. A second goal is to identify the in vivo niche(s) that promote parasexual reproduction. Our studies indicate that particular regions of the gastrointestinal tract may preferentially support phenotypi switching and parasexual reproduction. This possibility will be directly tested by the experiments outlined here, including quantification of each of the steps of the parasexual cycle in vivo. Together, completion of the proposed experiments will therefore determine where parasexual reproduction occurs in vivo, and the consequences of this program for generating recombinant strains with increased pathogenicity.